The overarching focus of my research is to understand how post-transcriptional regulation of gene expression, particularly at the level of mRNA metabolism, participates in modulating neuronal functions. It is now clear that synaptogenesis and synaptic plasticity rely on localized translation within dendrites of mRNAs encoding key synaptic structural and functional constituents. Yet, the molecular machinery underlying dendritic mRNAs localization and regulated expression is still poorly understood.
A primary subject of my past and current studies is the immediate early gene Arc, encoding a protein whose function is critical for LTP, LTD, memory consolidation, and homeostatic plasticity. Arc mRNA is also emerging as a unique example of how neuronal activity can control every known step of a mRNA’s life, ranging from its transcription, to its localization, stability and finally translation. In my previous work, I identified a novel pathway modulating Arc mRNA expression, which relies on the presence of introns in its 3’UTR. This unique genomic arrangement causes the mRNA to be targeted for destruction by the Nonsense Mediated Decay pathway (NMD), adding another degree of complexity to the already intricate journey of Arc mRNA expression at synapses.
To understand the underlying molecular mechanisms controlling Arc mRNA metabolism, I have been adopting parallel approaches aimed at the characterization of cis and trans-acting factors associated with its 3’UTR. On one hand, I am utilizing biochemical techniques to isolate the ribonucleoprotein particle associated with Arc mRNA 3’UTR in vivo. Mass spectrometry and deep sequencing of this complex will allow the identification of trans-acting factors and miRNAs whose binding is dependent on synaptic activity and splicing of the mRNA. In a parallel analysis, luciferase reporter constructs harboring deletions and mutations of Arc 3’UTR are being adopted.
This approach is aimed at understanding the molecular links between translational activation of Arc mRNA processing of its mRNA. Analyses are underway to test whether other dendritic mRNAs undergo similar regulatory pathways as Arc. Overall, the in vivo biochemical and functional characterization of both cis- and trans-acting factors controlling the expression of dendritic mRNAs, will allow a better understanding of the mechanisms controlling their expression, and likely shed light on key pathways that link dendritic mRNA localized expression to synaptic plasticity.